DNA-Binding Agent Trabectedin Combined With Recombinant Methioninase Is Synergistic to Decrease Fibrosarcoma Cell Viability and Induce Nuclear Fragmentation But Not Synergistic on Normal Fibroblasts.

BACKGROUND/AIM: The alkylating agent trabectedin, which binds the minor groove of DNA, is second-line therapy for soft-tissue sarcoma but has only moderate efficacy. The aim of the present study was to determine the synergistic efficacy of recombinant methioninase (rMETase) and trabectedin on fibrosarcoma cells in vitro, compared with normal fibroblasts. MATERIALS AND METHODS: HT1080 human fibrosarcoma cells expressing green fluorescent protein (GFP) in the nucleus and red fluorescent protein (RFP) in the cytoplasm and Hs27 normal human fibroblasts, were used. Each cell line was cultured in vitro and divided into four groups: no-treatment control; trabectedin treated; rMETase treated; and trabectedin plus rMETase treated. The dual-color HT1080 cells were used to quantitate nuclear fragmentation in each treatment group. RESULTS: The combination of rMETase and trabectedin was highly synergistic to decrease HT1080 cell viability. In contrast, there was no synergy on Hs27 cells. Moreover, nuclear fragmentation occurred synergistically with the combination of trabectedin and rMETase on dual-color HT1080 cells. CONCLUSION: The combination treatment of trabectedin plus rMETase was highly synergistic on fibrosarcoma cells in vitro suggesting that the combination can improve the outcome of trabectedin alone in future clinical studies. The lack of synergy of rMETase and trabectedin on normal fibroblasts suggests the combination is not toxic to normal cells. Synergy of the two drugs may be due to the high rate of nuclear fragmentation on treated HT1080 cells, and the late-S/G2 cell-cycle block of cancer cells by rMETase, which is a target for trabectedin. The results of the present study suggest the future clinical potential of the combination of rMETase and trabectedin for soft-tissue sarcoma.

The potential of methioninase for cancer treatment.

Cancer cells are addicted to L-methionine (L-Met) and have a much greater requirement for L-Met than normal cells due to excess transmethylation, termed the Hoffman effect. By targeting this vulnerability through dietary restriction of L-Met, researchers have been able to achieve promising results in inhibiting tumor growth and eradicating cancer cells. Methioninase (EC 4.4.1.11; METase) catalyzes the transformation of L-Met into α-ketobutyrate, ammonia, and methanethiol. The use of METase was initially limited due to its poor stability in vivo, high immunogenicity, and enzyme-induced inactivating antibodies. These issues could be partially resolved by PEGylation, encapsulation in erythrocytes, and various site-directed mutagenesis. The big breakthrough came when it was discovered that METase is effectively administered orally. The enzyme L-asparaginase is approved by the FDA for treatment of acute lymphoblastic leukemia. METase has more potential as a therapeutic since addiction to L-Met is a general and fundamental hallmark of cancer.

Roles of luxS in regulation of probiotic characteristics and inhibition of pathogens in Lacticaseibacillus paracasei S-NB.

Lactic acid bacteria (LAB) have excellent tolerance to the gastrointestinal environment and high adhesion ability to intestinal epithelial cells, which could be closely related to the LuxS/AI-2 Quorum sensing (QS) system. Here, the crucial enzymes involved in the synthesis of AI-2 was analyzed in Lacticaseibacillus paracasei S-NB, and the luxS deletion mutant was constructed by homologous recombination based on the Cre-lox system. Afterwards, the effect of luxS gene on the probiotic activities in L. paracasei S-NB was investigated. Notably, the tolerance of simulated gastrointestinal digestion, AI-2 production, ability of auto-aggregation and biofilm formation significantly decreased (p < 0.05 for all) in the S-NB△luxS mutant. Compared to the wild-type S-NB, the degree of reduction in the relative transcriptional level of the biofilm -related genes in Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923 was diminished when co-cultured with S-NB△luxS. Furthermore, the inhibitory effect of S-NB△luxS on the adhesion (competition, exclusion and displacement) of E. coli ATCC 25922 and S. aureus ATCC 25923 to Caco-2 cells markedly decreased. Therefore, comprehensive analysis of the role by luxS provides an insight into the LuxS/AI-2 QS system of L. paracasei S-NB in the regulation of strain characteristics and inhibition of pathogens.

Effects of luxS gene on growth characteristics, biofilm formation, and antimicrobial resistance of multi-antimicrobial-resistant Vibrio parahaemolyticus Vp2015094 isolated from shellfish.

AIMS: Vibrio parahaemolyticus is an important foodborne pathogen worldwide, which can cause gastroenteritis. This study aimed to investigate the effect of quorum sensing system LuxS/AI-2-related gene luxS on the biological characteristics and antimicrobial resistance of V. parahaemolyticus Vp2015094 from shellfish, which carried a multi-antimicrobial-resistant plasmid. METHODS AND RESULTS: The critical gene luxS related to the synthesis of AI-2 in V. parahaemolyticus Vp2015094 was knocked out by homologous recombination with suicide plasmid. The effect of luxS on the biological characteristics of V. parahaemolyticus was determined by comparing the growth, AI-2 activity, motility, biofilm formation ability, and antibiotic resistance between the wildtype strain and the luxS deletion mutant. Compared with wildtype strain, the production of AI-2, the motility and biofilm formation ability, antimicrobial resistance, and conjugation frequency of luxS deletion mutant strain were decreased. The transcriptome sequencing showed that the transcriptional levels of many genes related to motility, biofilm formation, antimicrobial resistance, and conjugation were significantly downregulated after luxS deletion. CONCLUSIONS: Quorum sensing system LuxS/AI-2-related gene luxS in V. parahaemolyticus Vp2015094 played an important role in growth characteristics, biofilm formation, antimicrobial resistance, and resistance genes' transfer.

Effects of metformin on Streptococcus suis LuxS/AI-2 quorum sensing system and biofilm formation.

Streptococcus suis (S. suis) regulates biofilm formation through LuxS/AI-2 quorum sensing system, increasing drug resistance and exacerbating infection. The anti-hyperglycaemic agent metformin has anti-bacterial and anti-biofilm activities. This study aimed to investigate the anti-biofilm and anti-quorum sensing activity of metformin in S. suis. We first determined the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of metformin on S. suis. The results indicated that metformin showed no obvious inhibitory or bactericidal effect. Crystal violet staining showed that metformin significantly inhibited the formation of S. suis biofilm at sub-MIC concentration, which was also confirmed by scanning electron microscopy. Then, we quantified the AI-2 signal molecules in S. suis, and the results showed that metformin had a significant inhibitory effect on the production of AI-2 signal in S. suis. Inhibition of enzyme activity and molecular docking experiments showed that metformin has a significant binding activity to LuxS protein. In addition, qRT-PCR results showed that metformin significantly down-regulated the expression of AI-2 synthesis-related genes luxS and pfs, and adhesion-related genes luxS, pfs, gapdh, sly, fbps, and ef. Western blotting also showed that metformin significantly reduced the expression of LuxS protein. Our study suggests that metformin seems to be a suitable candidate for the inhibition of S. suis LuxS/AI-2 QS system and prevention of biofilm formation, which provided a new idea for the prevention and control of S. suis.

Hydrogen sulfide alleviates mitochondrial damage and ferroptosis by regulating OPA3-NFS1 axis in doxorubicin-induced cardiotoxicity.

Ferroptosis is a major cause of cardiotoxicity induced by doxorubicin (DOX). Previous studies have shown that hydrogen sulfide (H2S) inhibits ferroptosis in cardiomyocytes and myoblasts, but the underlying mechanism has not been fully elucidated. In this study, we investigated the role of H2S in protecting against DOX-induced cardiotoxicity both in vivo and in vitro, and elucidated the potential mechanisms involved. We found that DOX downregulated the expression of glutathione peroxidase 4 (GPX4) and NFS1, and upregulated the expression of acyl-coenzyme A synthetase long-chain family member 4 (ACSL4) expression level, resulting in increased lipid peroxidation and ferroptosis. Additionally, DOX inhibited MFN2 expression and increased DRP1 and FIS1 expression, leading to abnormal mitochondrial structure and function. In contrast, exogenous H2S inhibited DOX-induced ferroptosis by restoring GPX4 and NFS1 expression, and reducing lipid peroxidation in H9C2 cells. This effect was similar to that of the ferroptosis antagonist ferrostatin-1 (Fer-1) in protecting against DOX-induced cardiotoxicity. We further demonstrated that the protective effect of H2S was mediated by the key mitochondrial membrane protein optic atrophy 3 (OPA3), which was downregulated by DOX and restored by exogenous H2S. Overexpression of OPA3 alleviated DOX-induced mitochondrial dysfunction and ferroptosis both in vivo and in vitro. Mechanistically, NFS1 has an inhibitory effect on ferroptosis, and NFS1 deficiency increases the susceptibility of cardiomyocytes to ferroptosis. OPA3 is involved in the regulation of ferroptosis by interacting with NFS1. Post-translationally, DOX promoted OPA3 ubiquitination, while exogenous H2S antagonized OPA3 ubiquitination by promoting OPA3 s-sulfhydration. In summary, our findings suggested that H2S protects against DOX-induced cardiotoxicity by inhibiting ferroptosis via targeting the OPA3-NFS1 axis. This provides a potential therapeutic strategy for the treatment of DOX-induced cardiotoxicity.

Recombinant Oral Methioninase (o-rMETase) Combined With Oxaliplatinum Plus 5-Fluorouracil Improves Survival of Mice With Massive Colon-Cancer Peritoneal Carcinomatosis.

BACKGROUND/AIM: The present study aimed to determine if oral methioninase (o-rMETase) combined with oxaliplatinum (OXA) plus 5-fluorouracil (5-FU) increases survival of mice with peritoneal-carcinomatosis formed from HCT-116 green fluorescent protein (GFP)-expressing colon-cancer cells implanted intra-peritoneally in nude mice. MATERIALS AND METHODS: HCT-116-GFP human colon-cancer cells (2×106) were injected intraperitoneally in athymic nude mice. Forty-five HCT-116-GFP colon-cancer peritoneal-carcinomatosis nude-mouse models were divided into the following groups: untreated control; combination of 5-FU (50 mg/kg, once a week), plus OXA (6 mg/kg, once a week); combination of 5-FU + OXA + o-rMETase (100 unit/day). Tumor growth was followed weekly using non-invasive GFP imaging for 3 weeks. At necropsy, tumor tissue was obtained. Frozen sections were made for fluorescence imaging. Tumor tissues were also stained with hematoxylin and eosin. The date of death of all mice was recorded. RESULTS: o-rMETase combined with 5-FU + OXA significantly reduced peritoneal growth of the HCT-116 tumor compared to the untreated control or the combination 5-FU and OXA group. Histological analysis revealed extensive necrosis induced by the o-rMETase + 5-FU + OXA combination. The combination of 5-FU plus OXA and o-rMETase achieved significantly longer survival of the mice with peritoneal carcinomatosis compared to the control or combination of 5-FU plus OXA treatments. CONCLUSION: o-rMETase shows future clinical promise for increasing the survival of patients with peritoneal metastasis of colon cancer when combined with first-line treatment of this recalcitrant disease.

Inhibitory Effect of Monoterpenoid Glycosides Extracts from Peony Seed Meal on Streptococcus suis LuxS/AI-2 Quorum Sensing System and Biofilm.

Streptococcus suis LuxS/AI-2 quorum sensing system regulates biofilm formation, resulting in increased pathogenicity and drug resistance, and diminished efficacy of antibiotic treatment. The remaining peony seed cake after oil extraction is rich in monoterpenoid glycosides, which can inhibit the formation of bacterial biofilm. In this study, we investigated the effect of seven major monocomponents (suffruticosol A, suffruticosol B, suffruticosol C, paeonifloin, albiflorin, trans-ε-viniferin, gnetin H) of peony seed meal on minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of S. suis. The results showed that the MICs of the seven single components were all greater than 200 µg/mL, with no significant bacteriostatic and bactericidal advantages. Crystal violet staining and scanning electron microscope observation showed that the seven single components had a certain inhibitory effect on the biofilm formation ability of S. suis at sub-MIC concentration. Among them, the ability of paeoniflorin to inhibit biofilm was significantly higher than that of the other six single components. AI-2 signaling molecules were detected by bioreporter strain Vibrio harvey BB170. The detection results of AI-2 signal molecules found that at 1/2 MIC concentration, paeoniflorin significantly inhibited the production of S. suis AI-2 signal, and the inhibitory effect was better than that of the other six single components. In addition, molecular docking analysis revealed that paeoniflorin had a significant binding activity with LuxS protein compared with the other six single components. The present study provides evidence that paeoniflorin plays a key role in the regulation of the inhibition of S. suis LuxS/AI-2 system and biofilm formation in peony seed meal.

Oral-recombinant Methioninase in Combination With Rapamycin Eradicates Osteosarcoma of the Breast in a Patient-derived Orthotopic Xenograft Mouse Model.

BACKGROUND/AIM: Primary osteosarcoma of the breast is a very rare malignancy that shares histological features with osteosarcoma. It is also highly sensitive to methionine restriction due to methionine addiction. We previously established a patient-derived orthotopic xenograft (PDOX) nude-mouse model derived from tumor tissue of a patient with primary mammary osteosarcoma. In the present study, we investigated the efficacy of oral-recombinant methioninase (o-rMETase), combined with rapamycin, an inhibitor of mammalian target of rapamycin (mTOR) kinase, on a mammary osteosarcoma PDOX nude-mouse model. MATERIALS AND METHODS: The PDOX mouse model was established by surgically transplanting a specimen of primary osteosarcoma of the breast into the mammary gland of nude mice. Mice implanted with tumors were randomly divided into four groups: Control group, N=5; rapamycin-treated group, N=5; o-rMETase-treated group, N=5; and a group treated with the combination of o-rMETase and rapamycin, N=5. Mice were treated for 2 weeks after transplantation, and tumor volume was measured during the treatment period. RESULTS: Treatment with the combination of rapamycin and o-rMETase eradicated the osteosarcoma of the breast compared to the untreated control (p=0.000008). o-rMETase alone did not significantly inhibit tumor growth, and rapamycin alone only partially inhibited the tumor (p=0.78 and p=0.018, respectively) compared to the untreated control. There was not a significant difference in mouse weight between the groups. CONCLUSION: The combination of rapamycin and o-rMETase was highly effective against primary osteosarcoma of the breast in a PDOX model, suggesting a future clinical strategy for this rare cancer type that currently has no first-line treatment.

Oral Installation of Recombinant Methioninase-producing Escherichia coli into the Microbiome Inhibits Colon-cancer Growth in a Syngeneic Mouse Model.

BACKGROUND/AIM: All cancer types so far tested are methionine-addicted. Targeting the methionine addiction of cancer with recombinant methioninase (rMETase) has shown great progress in vitro, in mouse models, and in the clinic. However, administration of rMETase requires multiple doses per day. In the present study, we determined if rMETase-producing Escherichia coli JM109 (E. coli JM109-rMETase) might be an effective anticancer agent when installed into the microbiome. MATERIALS AND METHODS: E. coli JM109-rMETase was administered to a syngeneic model of MC38 colon cancer growing subcutaneously in C57BL/6 mice. JM109-rMETase was administered orally by gavage to the mice twice per day. Tumor size was measured with calipers. RESULTS: The administration of E. coli JM109-rMETase twice a day significantly inhibited MC38 colon-cancer growth. E. coli JM109-rMETase was found in the stool of treated mice, indicating it had entered the microbiome. CONCLUSION: The present study indicates the potential of microbiome-based treatment of cancer targeting methionine addiction.

Extent and Instability of Trimethylation of Histone H3 Lysine Increases With Degree of Malignancy and Methionine Addiction.

BACKGROUND/AIM: Methionine addiction is a fundamental and general hallmark of cancer, termed the Hoffman effect. Methionine addiction is due to excessive use of and dependence on methionine by cancer cells. In the present report, we correlated the extent of methionine addiction and degree of malignancy with the amount and stability of methylated histone H3 lysine marks. MATERIALS AND METHODS: We established low- and high-malignancy variants from a parental human pancreatic-cancer cell line and compared their sensitivity to methionine restriction and histone H3 lysine methylation status. RESULTS: A low-malignancy, low-methionine-addiction revertant of the parental pancreatic-cancer cell line had less methylated H3K9me3 and was less sensitive to methionine restriction effected by recombinant methioninase (rMETase) than the parental cell line. A high-malignancy variant of the pancreatic cancer cell line had increased methylated H3K9me3 and was more sensitive to methionine restriction by rMETase with regard to inhibition of proliferation and to instability of histone H3 lysine methylation than the parental cell line. Orthotopic malignancy in nude mice was reduced in the low-methionine-addiction revertant and greater in the high-malignancy variant than in the parental cell line. CONCLUSION: The present study indicates that the degree of malignancy is linked to the extent of methionine addiction and the level and instability of trimethylation of histone H3, suggesting these phenomena are linked as a fundamental basis of oncogenic transformation.

Paeoniflorin reduce luxS/AI-2 system-controlled biofilm formation and virulence in Streptococcus suis.

Streptococcus suis (S. suis), more specifically serotype 2, is a bacterial pathogen that threatens the lives of pigs and humans. Like many other pathogens, S. suis exhibits quorum sensing (QS) system-controlled virulence factors, such as biofilm formation that complicates treatment. Therefore, impairing the QS involving LuxS/AI-2 cycle in S. suis, may be a promising alternative strategy for overcoming S. suis infections. In this study, we investigated paeoniflorin (PF), a monoterpenoid glycoside compound extracted from peony, as an inhibitor of S. suis LuxS/AI-2 system. At a sub-minimal inhibitory concentration (MIC) (1/16 MIC; 25 µg/ml), PF significantly reduced biofilm formation by S. suis through inhibition of extracellular polysaccharide (EPS) production, without affecting bacterial growth. Moreover, evidence was brought that PF reduces AI-2 activity in S. suis biofilm. Molecular docking indicated that LuxS may be the target of PF. Monitoring LuxS enzymatic activity confirmed that PF had a partial inhibitory effect. Finally, we showed that the use of PF in a mouse model can relieve S. suis infections. This study highlighted the anti-biofilm potential of PF against S. suis, and brought evidence that it may as an inhibitor of the LuxS/AI-2 system to prevent S. suis biofilm-related infections. PF can thus be used as a new type of natural biofilm inhibitor for clinical application.

Conjugates of methionine γ-lyase with polysialic acid: Two approaches to antitumor therapy.

The methionine dependence is a well known phenomenon in metabolism of cancer cells. Methionine γ-lyase (EC 4.4.1.11, MGL) catalyzes the γ-elimination reaction of L-methionine and thus could effectively inhibit the growth of malignant cells. Recently we have demonstrated that the mutant form of the enzyme C115H MGL can be used as a component of the pharmacological pair enzyme/S-(allyl/alkyl)-L-cysteine sulfoxides to yield thiosulfinates in situ. Thiosulfinates were shown to be toxic to various cancer cell lines. Therefore the application of the enzyme in enzyme pro-drug therapy may be promising. The conjugates of MGL and C115H MGL with polysialic acid were obtained and their kinetic and pharmacokinetic parameters were determined. The formation of polysialic shell around the enzyme was confirmed by atomic force microscopy. The half-life of conjugated enzymes increased 3-6 times compared to the native enzyme. The cytotoxic effect of conjugated MGL against methionine dependent cancer cell lines was increased two times compared to the values for the native enzymes. The anticancer efficiency of thiosulfinates produced by pharmacological pair C115H MGL/S-(allyl/alkyl)-L-cysteine sulfoxides was demonstrated in vitro. The results indicate that the conjugates of MGL with polysialic acid could be new antitumor drugs.

Oral recombinant methioninase combined with paclitaxel arrests recalcitrant ovarian clear cell carcinoma growth in a patient-derived orthotopic xenograft (PDOX) nude-mouse model.

PURPOSE: Advanced ovarian clear cell carcinoma (OCCC) is a recalcitrant disease, often resistant to the first-line platinum-based therapy. Using a novel patient-derived orthotopic xenograft (PDOX) nude-mouse model of OCCC, we tested whether oral-recombinant methioninase (o-rMETase) could enhance the efficacy of paclitaxel (PTX). METHODS: The OCCC PDOX model was established and passaged in nude mice. The OCCC PDOX models were randomized into 5 groups. G1: untreated control; G2: paclitaxel (PTX) (20 mg/kg, intraperitoneal (i.p.) injection, weekly); G3: o-rMETase (100 units, oral, daily); G4: PTX (20 mg/kg, i.p. injection, weekly) + carboplatinum (CBDCA) (40 mg/kg, i.p. injection weekly); G5: PTX (20 mg/kg, i.p. injection, weekly) + o-rMETase (100 units, oral, daily). The treatment period was 2 weeks. RESULTS: The combination of PTX and o-rMETase arrested OCCC tumor growth (relative tumor volume: 1.09 ± 0.63 (mean ± SD)) compared with the untreated control (relative tumor volume: 3.92 ± 1.04 (mean ± SD)) (p < 0.0001). There was no significant difference in relative tumor volume between PTX plus o-rMETase and PTX plus CBDCA (relative tumor volume: 1.39 ± 0.37 (mean ± SD)) (p = 0.93). CONCLUSION: PTX plus o-rMETase arrested the OCCC tumor growth. o-rMETase is readily administered and can greatly enhance first-line therapy of a recalcitrant cancer. The novel and effective treatment strategy in the present report has future clinical potential for patients with OCCC, especially for patients who cannot well tolerate platinum-based therapy.

Triple-Methyl Blockade With Recombinant Methioninase, Cycloleucine, and Azacitidine Arrests a Pancreatic Cancer Patient-Derived Orthotopic Xenograft Model.

OBJECTIVES: Methionine addiction is a fundamental and general hallmark of cancer caused by enhanced methyl flux. In the present study, we effected a novel methionine-methylation blockade to target a patient-derived orthotopic xenograft model of pancreatic cancer. METHODS: The pancreatic cancer patient-derived orthotopic xenograft mouse models were randomized into 6 groups of 8 mice each and treated for 2 weeks: untreated control; azacitidine; oral recombinant methioninase (o-rMETase); o-rMETase plus cycloleucine; o-rMETase plus cycloleucine plus azacitidine (triple-methyl blockade therapy); and gemcitabine (positive control). RESULTS: Triple-methyl blockade therapy arrested tumor growth (mean relative tumor volume, 1.03 [standard deviation, 0.36]) and was significantly more effective compared with azacitidine (P = 0.0001); o-rMETase (P = 0.007); or o-rMETase plus cycloleucine (P = 0.04). Gemcitabine alone also inhibited but did not arrest tumor growth (mean relative tumor volume, 1.50 [standard deviation, 0.30]). The percentage of cancer cells that were negative for 5-methylcytosine staining in immunohistochemistry, indicating reduction of DNA methylation, increased with triple-methyl blockade therapy (37.5%), compared with gemcitabine (1.8%); o-rMETase (2.8%); azacitidine (9.0%); or o-rMETase plus cycloleucine (10.6%). CONCLUSIONS: This new concept of triple-methyl blockade therapy has clinical potential for pancreatic cancer, which is currently a recalcitrant disease.

Preparation of Internalizing RGD-Modified Recombinant Methioninase Exosome Active Targeting Vector and Antitumor Effect Evaluation.

BACKGROUND/AIMS: Targeted drug delivery vehicles with low immunogenicity and toxicity are needed for cancer therapy. Here, we prepare an active targeting drug carrier of low immunogenicity and toxicity for targeted therapy. METHODS: Immature dendritic cells (imDCs) from BALB/c mice were used as donor cells of exosomes (Exos) that were transfected with the plasmids expressing fusion proteins of a tumor-targeting peptide known as internalizing RGD (iRGD) to construct a type of tumor-targeting iRGD-Exos and observe the interaction between these iRGD-Exos. Also, recombinant methioninase (rMETase) was loaded into the iRGD-Exos by electroporation to construct iRGD-Exos-rMETase and to assess the tumor-targeting function of the iRGD-Exos-rMETase. Finally, 30 BALB/c were randomly divided into five groups (n = 6), to observe tumor growth in vivo. RESULTS: The iRGD-Exos-rMETase was 99.58 nm in diameter and presented a unique "goblet" structure under transmission electron microscopy (TEM), with the encapsulation efficiency (EE) of 19.05%. iRGD-Exos-rMETase group has the strongest tumor suppressive effect. Compared to the iRGD-Exos-rMETase group, rMETase group and the blank-Exos-rMETase group were less effective, while the PBS group and the iRGD-Exos group showed no inhibitory effect on tumor growth. After treatment, the iRGD-Exos-rMETase group had gastric tumors significantly smaller and lighter than the other groups (P < 0.05). CONCLUSION: The iRGD-Exos-rMETase is an effective antitumor therapy that delivers rMETase to tumor tissue using the iRGD-Exos. With its favorable inhibitory effect and tumor-targeting function, the iRGD-Exos-rMETase shows excellent potential value and exciting prospects in clinical applications.

Oral Recombinant Methioninase Sensitizes a Bladder Cancer Orthotopic Xenograft Mouse Model to Low-dose Cisplatinum and Prevents Metastasis.

BACKGROUND/AIM: The aim of the study was to determine if oral recombinant methioninase (o-rMETase) can sensitize an orthotopic bladder tumor in nude mice to low-dose cisplatinum (CDDP). MATERIALS AND METHODS: The green fluorescent protein (GFP)-expressing UM-UC-3-GFP bladder cancer was surgically orthotopically implanted (SOI) to the bladder in nude mice. The treatment was initiated when the primary tumor volume reached 100 mm3 Mice were assigned to 3 groups: G1: Saline vehicle (0.1 ml per mouse, oral, twice per day); G2: low-dose CDDP (0.5 mg/kg, intraperitoneal twice per week); G3: o-rMETase + low-dose CDDP (100 units per mouse, oral, twice per day + 0.5 mg/kg, intraperitoneal twice per week, respectively). Tumor volume and body weight were measured twice per week. The expression of Ki-67 was detected by immunohistochemistry to evaluate cell proliferation. RESULTS: The combination of o-rMETase and low-dose CDDP increased inhibition efficacy compared to low-dose CDDP monotherapy, on primary-tumor growth (p=0.032) and metastasis (p=0.002). CONCLUSION: The combination of o-rMETase with low-dose CDDP has future clinical potential for bladder cancer.

A Randomized, Double-Blind Study Comparing Pharmacokinetics and Pharmacodynamics of Proposed Biosimilar ABP 798 With Rituximab Reference Product in Subjects With Moderate to Severe Rheumatoid Arthritis.

ABP 798 is a proposed biosimilar to rituximab reference product (RP), an anti-CD20 monoclonal antibody. Pharmacokinetics (PK), pharmacodynamics (PD), and safety results from the comparative clinical study that evaluated the PK, PD, safety, efficacy, and immunogenicity of ABP 798 versus rituximab RP are presented here. Subjects with moderate to severe rheumatoid arthritis (RA) received 2 doses of ABP 798, United States-sourced RP (rituximab US) or European Union-sourced RP (rituximab EU), each consisting of two 1000-mg infusions 2 weeks apart. For the second dose (week 24), ABP 798- and rituximab EU-treated subjects received the same treatment; rituximab US-treated subjects transitioned to ABP 798. End points included area under the serum concentration-time curve from time 0 extrapolated to infinity and maximum observed serum concentration following the second infusion of the first dose (PK) and percentage of subjects with complete CD19+ cell depletion days 1-33 (PD). Primary analysis established PK similarity between ABP 798 and rituximab RP based on 90% confidence intervals of the adjusted geometric mean ratios being within a prespecified equivalence margin of 0.8 and 1.25. Complete CD19+ B-cell depletion on day 3 among groups confirmed PD similarity. These findings demonstrated PK/PD similarity between ABP 798 and rituximab RP in subjects with moderate to severe RA.

Recombinant Methioninase Combined With Tumor-targeting Salmonella typhimurium A1-R Induced Regression in a PDOX Mouse Model of Doxorubicin-resistant Dedifferentiated Liposarcoma.

BACKGROUND/AIM: Dedifferentiated liposarcoma (DDLPS) is associated with a poor survival rate even with multi-modality treatment. In the present study, we evaluated the efficacy of recombinant methioninase (rMETase) combined with tumor-targeting Salmonella typhimurium (S. typhimurium) A1-R against a doxorubicin-resistant DDLPS in a patient-derived orthotopic xenograft (PDOX) mouse model. MATERIALS AND METHODS: A recurrent high-grade DDLPS from the right retroperitoneum of a patient was grown orthotopically in the retroperitoneum of nude mice to establish a PDOX model. The PDOX models were randomly divided into the following groups: Control, no treatment; doxorubicin monotherapy; rMETase monotherapy; S. typhimurium A1-R monotherapy; S. typhimurium A1-R and rMETase combination therapy. Tumor length and width were measured before and after treatment. RESULTS: On day 14 after treatment, all treatments significantly inhibited DDLPS PDOX tumor growth compared to the untreated control except for doxorubicin monotherapy. rMETase combined with S. typhimurium A1-R was significantly more effective and regressed tumor volume compared to either rMETase or S. typhimurium A1-R alone. The relative body weight did not significantly differ between days 0 and 14 for individual groups. CONCLUSION: The combination of rMETase and S. typhimurium A1-R has important clinical potential for this recalcitrant sarcoma.

Oral dosing of Recombinant Methioninase Is Associated With a 70% Drop in PSA in a Patient With Bone-metastatic Prostate Cancer and 50% Reduction in Circulating Methionine in a High-stage Ovarian Cancer Patient.

BACKGROUND/AIM: Methionine addiction is a general and fundamental hallmark of cancer. Methionine addiction can be targeted by methionine restriction (MR). Our laboratory has studied methionine addiction in cancer and MR for almost 50 years. The present study describes oral recombinant methioninase (o-rMETase), as a supplement, to induce MR in cancer patients. PATIENTS AND METHODS: One patient, a 67-year-old female with high-stage ovarian cancer took o-rMETase twice a day at 250 units per dose for approximately one month. A second patient, a 76-year-old male with bone-metastatic prostate cancer, took o-rMETase twice a day at 250 units per dose during three months. RESULTS: The first patient's circulating methionine levels decreased 50% within 4 hours of taking 250 units of o-rMETase. The second patient's PSA dropped approximately 70% over 3 months. During this time the patient's hemoglobin increased. CONCLUSION: o-rMETase has no side effects and is potentially efficacious. Future studies involving larger cohorts of patients with high-stage cancer are required to determine if o-rMETase, as a supplement, can increase survival and improve the quality of life.